High-affinity Ca(2+)-activated ATPases that do not show any demonstrable dependence on Mg2+ have been reported in the plasma membranes of different trypanosomatids, and it has been suggested [McLaughlin (1985) Mol. Biochem. Parasitol. 15, 189-201; Ghosh, Ray, Sarkar & Bhaduri (1990) J. Biol. Chem. 265, 11345-11351] that these enzymes may have a role in Ca2+ transport by the plasma membrane and in the regulation of intracellular Ca2+ in these parasites. In this report we investigated Ca2+ transport by Trypanosoma cruzi plasma membrane vesicles using Arsenazo III as a Ca2+ indicator. These vesicles accumulated Ca2+ upon addition of ATP only when Mg2+ was present and released it in response to the Ca2+ ionophore A23187, but were insensitive to inositol 1,4,5-trisphosphate. Ca2+ transport was insensitive to antimycin A, oligomycin and carbonyl cyanide p-trifluorophenylhydrazone, ruling out any mitochondrial contamination. Staurosporine and phorbol myristate acetate had no effect on this activity, while low concentrations of vanadate (10 microM) completely inhibited it. In addition, we describe a high-affinity vanadate-sensitive (Ca(2+)-Mg2+)-ATPase in the highly enriched plasma membrane fraction of T. cruzi. Kinetic studies indicated that the apparent Km for free Ca2+ was 0.3 microM. On the other hand, Ca(2+)-ATPase activity and Ca2+ transport were both stimulated by bovine brain calmodulin and by endogenous calmodulin purified from these cells. In addition, trifluoperazine and calmidazolium, at concentrations in the range in which they normally exert anti-calmodulin effects, inhibited the calmodulin-stimulated Ca(2+)-ATPase activity. These observations support the notion that a Mg(2+)-dependent plasma membrane Ca2+ pump is present in these parasites.

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In areas of low transmission of schistosomiasis, the evaluation of the success of control depends on reliable diagnostic tests. Under such conditions, some of the serological tests better estimate the real prevalence of this parasitosis than the classical stool examinations. On the search of highly sensitive and specific antigenic fractions for use in serological tests, an immunoblot technique with a luminescent substrate was used in order to evaluate, under dissociating and reducing conditions, the Schistosoma mansoni adult worm antigen (AWA). The sera of 30 infected Venezuelan children were assayed for specific recognition of AWA by IgG, IgM, IgE, IgA, and the four IgG subclasses. Eight highly specific polypeptide molecules from the parasite of 118, 114, 105, 74, 71, 45, 36, and 30 kDa were recognized by total IgG. Additionally, IgG1 and IgG2 recognized a molecule of 100 kDa and IgM one of 77 kDa. The present data suggests that certain molecules from the adult worm, specially the 36 kDa, might be relevant in the specific immunodiagnosis of this parasitic disease. The fact that the children antibodies were able to recognize the primary structure of these antigens, will allow us to chemically synthesize the relevant linear epitopes.

Oral transmission of Trypanosoma cruzi is a frequent cause of acute Chagas disease (ChD). In the present cross-sectional study, we report the epidemiological, clinical, serological and molecular outcomes of the second largest outbreak of oral ChD described in the literature. It occurred in March 2009 in Chichiriviche de la Costa, a rural seashore community at the central littoral in Venezuela. The vehicle was an artisanal guava juice prepared at the local school and Panstrongylus geniculatus was the vector involved. TcI genotype was isolated from patients and vector; some showed a mixture of haplotypes. Using molecular markers, parasitic loads were high. Eighty-nine cases were diagnosed, the majority (87.5%) in school children 6–15 years of age. Frequency of symptomatic patients was high (89.9%) with long-standing fever in 87.5%; 82.3% had pericardial effusion detected by echocardiogram and 41% had EKG abnormalities. Three children, a pregnant woman and her stillborn child died (5.6% mortality). The community was addressed by simultaneous determination of specific IgG and IgM, confirmed with indirect hemagglutination and lytic antibodies. Determination of IgG and IgA in saliva had low sensitivity. No individual parasitological or serological technique diagnosed 100% of cases. Culture and PCR detected T. cruzi in 95.5% of examined individuals. Based on the increasing incidence of oral acute cases of ChD, it appears that food is becoming one of the most important modes of transmission in the Amazon, Caribbean and Andes regions of America.

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